This invention is related to synthesis of activated iodo derivatives and their use as antineoplastic and antiviral agents by targeting the zinc finger regions of metalloregulatory proteins such as p-ADPRT and nucleocapsid of HIV. A series of activated iodo-benzamide derivatives are described as antineoplastic and antiviral drug compounds. The compounds generally possess a chelating group, a thiol trapping group and an activating group. The presumptive mechanism of action in preventing cancer cell and virus replication is through inhibition of the binding of transcription factors to zinc finger binding domains. The compounds are effective in inhibiting growth of a variety of human and animal tumor and leukemia cell lines at low doses.
Zinc Finger Proteins
In the past several years, a series of discoveries revealed that several proteins contain metal ions, particularly zinc ions (Zn.sup.2+), that play fundamental roles in stabilizing specific protein conformations (Berg, J. M., J. Bioj. Chem., 265: 6513-6516, 1990; Berg, J. M., In Progress in Inorganic chemistry, 37: 143-190, 1989). Many of these metalloproteins are involved in nucleic acid binding and in gene regulation (Bravo, R., Cell Growth and Differentiation, 1: 305-309, 1990; Evans; R. M., and Hollenberg, S. M., Cell, 52: 1-3, 198).
The nuclear eukaryotic enzyme poly(ADP-ribose)polymerase [PARP (EC 2.4.2.30)] catalyzes the transfer of the ADP-ribose moiety of nictotinamide adenine dinucleotide (NAD+) to nuclear aceptor proteins, in response to DNA strand-break formation (Udea et al., Annual Review of Biochemistry, 54, 73, 1985; Boulikas, T., Toxicology Letters, 67, 129, 1993; De Murcia et al., Trends in Biochemical Sciences, 172, 1994). The protein-bound linear and branched-chain homo-ADP polymers thus formed are implicated in a number of important cellular processes, including:
1. DNA repair--Shell, S., Advances in Radiation Biology, 11, 1, 1984; Clear, J. E. et al., Mutation Research, 257, 1, 1991. PA1 2. Cellular differentiation--Lautier et al., Molecular and Cellular Biochemistry, 122, 171, 1993. PA1 3. Gene expression--Boulikas, T., Toxicology Letters, 67, 129, 1993. PA1 4. Apoptosis--Kaufmann et al., Cancer Research, 53, 3976, 1993. PA1 1. A group acting as a competing chelating agent or sequestration agent. PA1 2. A functional group or groups to trap the released sulfide moiety. PA1 3. A functional group or groups to activate the functional trapping group. PA1 4. Conformation of the molecule to facilitate receptor/ligand interaction ##STR11## PA1 1. The chelating group at the amide linkage sequesters the zinc ion from the zinc fingers. PA1 2. As soon as the Zn ion is depleted from the system, the free thiol group generated will react with the functional group at the ortho position of the molecule forming a stable non-reversible linkage. The iodo group is selected for this purpose. The nucleophilic sulfide ion displaces the iodo group. PA1 3. The displacement of the iodo group is facilitated by the activating group, namely an electron withdrawing group. For our purpose we have selected one or two nitro or groups.
The human enzyme (116 kDa) is multifunctional and comprises an N-terminal DNA binding (46 kDa) containing two zinc fingers, a central automodification site (22 kDa) and a C-terminal domain (54 kDa).
Apoptosis and ADPRT
Apoptosis, or programmed cell death, plays an essential role in specific cell deletion during normal embryonal and adult development. Apoptotic cells are characterized by fragmentation of nuclear DNA and formation of apoptotic bodies. Molecular genetic analysis has revealed the involvement of several deaths and survival genes that are regulated by extracellular and intracellular factors. There are multiple inducers and inhibitors which interact with target cell specific receptors and transduce signals involved in cellular proliferation, cell cycle progression and programmed cell death. The elimination of tumor cell populations by applying lethal doses of chemotherapeutic agents or radiation is a well-established strategy in cancer therapy. Recent discoveries in the field of apoptotic cell death promise to have a significant impact on antitumor therapies. Apoptosis is known to be an active process which can be artificially manipulated by several molecular pathways.
Poly ADPRT has been consistently linked to the DNA repair process. ADP ribosylation levels have been mechanistically associated to human disease after activation of poly ADPRT by DNA damage by external sources. Firstly, poly ADPRT activity is dose-dependently up-regulated by reduced glutathione and down regulated by oxidized glutathione which establishes redox regulation of the enzyme. Secondly, the two zinc fingers in the two DNA binding domain of the poly ADPRT gene cysteine residues which, if oxidized, would presumably prevent DNA binding and participation in DNA repair.
The use of aromatic C-nitroso derivatives has been described in U.S. Pat. No. 5,516,941 for the treatment of diseases caused by viruses. The chemotherapeutic activity of an iodonitro derivative has been reported by Mendeleyev (Biochemical Pharmacology, 50, 705-714, 1995). The importance of the formation of a nitroso group for the compound's activity was disclosed in U.S. Pat. No. 5,670,518. The disclosed data were obtained with reduced glutathione thus indicating that the existence of free sulfide ion seems to be important for the activity of the reported compounds. Zinc fingers involve sulfide ions in the co-ordination of the metal ion, namely Zn.sup.2+. In the disclosed zinc ejection experiments, EDTA was used as one of the reagents and no data were provided in the absence of EDTA. Disulfide substituted benzamide has been shown as a zinc finger inhibitor (Turfin et al., Science, 270, 1194, 1995).
The compounds described by this invention have combined and enhanced functionalties which inhibit cancer cell and virus replication by binding to and disrupting zinc finger binding domains. They exhibit high efficacy in inhibiting cancer cell growth in vitro.